Regulation of galectin-9 expression and release in Jurkat T cell line cells

Ecalectin/galectin-9 was recently described as a novel eosinophil chemoattractant highly expressed in immune tissues. We investigated the regulation of galectin-9 expression and release in Jurkat (a T cell line) cells. We demonstrated that medium and long-sized galectin-9 isoforms were constitutively expressed, and phorbol 12-myriastate 13-acetate (PMA) upregulated the level of galectin-9 mRNA in Jurkat cells. Western blotting and flow cytometry analyses revealed that PMA stimulation resulted in the upregulation of both intracellular and surface galectin-9 protein. The stimulated Jurkat cells simultaneously released evident eosinophil chemoattractant activity (ECA). Main ECA was adsorbed by both lactose and anti-galectin-9 antibody affinity column, suggesting that the ECA was ascribed to galectin-9. When Jurkat cells were stimulated with PMA in the presence of a BB94, a matrix metalloproteinase (MMP) inhibitor, but not tissue inhibitor of metalloproteinase-1 (TIMP-1), the release of galectin-9 was suppressed in a dose-dependent manner. We further found that calphostin c, a protein kinase c (PKC) inhibitor, weakly but significantly suppressed the release of galectin-9. The present data suggested that galectin-9 production in Jurkat cells is provoked by the stimulation with PMA and that some MMP and PKC is, at least, partly involved in the release of galectin-9 from Jurkat cells.     

Progress in arylpiperazine synthesis by the catalytic amination reaction

Careful base and solvent optimization for catalytic amination is described. A Pd-catalyzed amination between some arylbromide and unprotected piperazine (1 equiv) was efficiently carried out with Pd/BINAP catalyst in a toluene–DBU solvent system, which is useful for the one-pot preparation of unsymmetrical piperazine through amination and in-situ N-protection. Reaction with N-BOC-piperazine was also successful in toluene–DBU or more polar NMP with Cs₂CO₃ as a key base. No reports have previously reported such solvent and base optimization in arylpiperazine synthesis.     

Structural and drug-binding properties of alpha(1)-acid glycoprotein in reverse micelles

Alpha(1)-acid glycoprotein (AGP) is a glycoprotein that consists of 183 amino acid residues and five carbohydrate chains and binds to neutral and basic drugs. We examined the structural properties and ligand-binding capacity of AGP in interactions with reverse micelles. Also, detailed information was obtained by comparing several different states of AGP. Interaction with reverse micelles induced a unique conformational transition (beta-sheet to alpha-helices) in AGP and decreased the binding capacity for the basic drug, chlorpromazine and the steroid hormone, progesterone to AGP. These structural conformations are very similar to those observed under conditions of acidity and high ionic strength (pH 2.0, 1.5 M NaCl). This structure seems to be an intermediate between the native state and the denatured state, possibly a molten globule. The present results suggest that when AGP interacts with the biomembrane, it undergoes a structural transition to a unique structure that differs from the native and denatured states and has a reduced ligand-binding capacity.     

TAK1-TAB1 fusion protein: a novel constitutively active mitogen-activated protein kinase kinase kinase that stimulates AP-1 and NF-kappaB signaling pathways

TAK1 mitogen-activated protein kinase kinase kinase (MAP3K) is activated by its specific activator, TAK1-binding protein 1 (TAB1). A constitutively active TAK1 mutant has not yet been generated due to the indispensable requirement of TAB1 for TAK1 kinase activity. In this study, we generated a novel constitutively active TAK1 by fusing its kinase domain to the minimal TAK1-activation domain of TAB1. Co-immunoprecipitation assay demonstrated that these domains interacted intra-molecularly. The TAK1-TAB1 fusion protein showed a significant MAP3K activity in vitro and activated c-Jun N-terminal kinase/p38 MAPKs and IkappaB kinase in vivo, which was followed by increased production of interleukin-6. These results indicate that the fusion protein is useful for characterizing the physiological roles of the TAK1-TAB1 complex.     

DNA aqueous solution used for dialytical removal and enrichment of dioxin derivatives

In the present study, a dialytic method that uses a DNA aqueous solution to remove and enrich dioxins from polluted water was proposed. Circular dichroism (CD) and fluorescent spectra indicated that dibenzo-p-dioxin (DD), dibenzofuran (DF) and biphenyl (BP), which are dioxin derivatives, form complexes with DNA. Their experimental dialytic sorption coefficients were measured by quantifying the concentrations of DD, DF, and BP in aqueous solutions before and after dialysis of the DNA solution, and the values were 2.1×10⁵, 1.3×10⁵, and 1.5×10⁷, respectively. As a simulated water treatment model, DNA solution was dialyzed in an aqueous mixture of DD, DF, and BP for 96 h, the HPLC studies showed that the dioxin derivatives have been concentrated in the DNA solution about 200 times. The dialyzed DNA solution was reusable by an extraction with hexane.     

Pleiotropic phenotypes of fission yeast defective in ubiquinone-10 production. A study from the abc1Sp (coq8Sp) mutant

We previously constructed two Schizosaccahromyces pombe ubiquinone-10 (or Coenzyme Q10) less mutants, which are either defective for decaprenyl diphosphate synthase or p-hydroxybenzoate polyprenyl diphosphate transferase. To further confirm the roles of ubiquinone in S. pombe, we examined the phenotype of the abc1Sp (coq8Sp) mutant, which is highly speculated to be defective in ubiquinone biosynthesis. We show here that the abc1Sp defective strain did not produce UQ-10 and could not grow on minimal medium. The abc1Sp-deficient strain required supplementation with antioxidants such as cysteine or glutathione to grow on minimal medium. In support of the antioxidant function of ubiquinone, the abc1Sp-deficient strain is sensitive to H2O2 and Cu2+. In addition, expression of the stress inducible ctt1 gene was much induced in the ubiquinone less mutant than wild type. Interestingly, we also found that the abc1-deficient strain as well as other ubiquinone less mutants produced a significant amount of H2S, which suggests that oxidation of sulfide by ubiquinone may be an important pathway for sulfur metabolism in S. pombe. Thus, analysis of the phenotypes of S. pombe ubiquinone less mutants clearly demonstrate that ubiquinone has multiple functions in the cell apart from being an integral component of the electron transfer system.     

Neonatal presentation of adult-onset type II citrullinemia

Adult-onset type II citrullinemia (CTLN2) is characterized by a liver-specific argininosuccinate synthetase deficiency caused by a deficiency of the citrin protein encoded by the SLC25A13 gene. Until now, however, no SLC25A13 mutations have been reported in children with liver diseases. We described three infants who presented as neonates with intrahepatic cholestasis associated with hypermethioninemia or hypergalactosemia detected by neonatal mass screening. DNA analyses of SLC25A13 revealed that one patient was a compound heterozygote for the 851de14 and IVS11+IG-->A mutations and two patients (siblings) were homozygotes for the IVS11+lG-->A mutation. These results suggested that there may be a variety of liver diseases related to CTLN2 in children.